Previous work in our laboratory has shown the presence of a factor (or factors) in plasma of saline loaded dogs which inhibits toad bladder sodium transport. Biochemical and physiologic characterization of the factor is being carried out. Extracts of plasma are prepared on Biogel P 2 in 1.0 M acetic acid. The active fraction is chromatographed by high pressure liquid chromatography (HPLC) on a cation exchange column in pyridine acetate. Peptide and amino acid peaks are detected by monitoring for fluorescamine derivatives. Fluorescamine is a fluorophore which reacts directly with primary amines. Biologic activity is present in the void volume. The active fraction on reverse phase chromatography contains a unique peak not present in inactive fractions. These studies suggest the toad bladder inhibitor is a small peptide, and provide the basis for development of a chemical assay for the factor. A natriuretic factor in plasma of volume expanded dogs and rats has also been demonstrated. This factor is different from the bladder inhibitor. Biologic activity, molecular weight, and behavior on Biogel P 2 and ion exchange chromatography are different. A possible biochemical relationship between the two factors is under investigation.